Motivated by the limitations of current synthesis methods for long (30-70 amino acids) peptides, the increasing importance of peptide drugs, and the clear need for low-cost expression and easy purification of peptides, the goal of this Phase I research proposal is to develop the recombinant expression and non-chromatographic purification of functionally active peptides at research-scale quantities at high purity levels, high yield, low cost and at similar biological activity to peptides produced by other chemical or biosynthetic methods, but with significantly lower cost. The enabling technology that will allow Phase Bioscience to achieve this objective is the delta Phase(TM) System, a novel platform technology for high-level expression and simple, non-chromatographic purification using a class of polypeptide tags, termed elastin- like polypeptides (ELPs). The fundamental basis of this technology is the discovery that proteins and peptides fused to an ELP retain the environmentally triggered (temperature, salt) soluble-insoluble phase transion behavior of ELPs, and that this reversible phase transition can be carried out in cell lysate to separate ELP fusions from other cellular proteins to homogeneity by a few phase transition cycles. The impact of the technology will be as follows: Development of Novel Medicines. As unique pharmaceutical targets are identified through functional genomics, they will need to be expressed and purified for evaluation (target validation) and screening (high- throughput screening and X-ray crystallography). The PhaseBio technology will rapidly expand the pool of target peptides by 1) enabling first-time, cost-effective expression of recombinant peptides; 2) facilitating the evaluation of peptide variants by enabling the production of up to 96 peptide variants simultaneously in only a matter of weeks; 3) eliminating the need to spend long development time looking for "shorter" active peptides due to manufacturing challenges and costs; and 4) enabling the rapid scale-up of a simple 4-step manufacturing process. The Specific Aims of the project include: 1. Express five pharmaceutically-relevant recombinant fusion peptides; 2. Purify the peptides to at least 90% purity, as measured by LC-MS. 3. Demonstrate similar bioactivity of peptides produced via Phase Bio's technology compared to the same peptides produced by commercially available chemical synthesis or recombinant peptide manufacturers. [unreadable] [unreadable] [unreadable]